Enteric infection with an obligate intracellular parasite, Encephalitozoon cuniculi, in an experimental model.

Rabbits were intrarectally infected with 3 doses (5 x 10(3), 5 x 10(5), and 5 x 10(7] of an obligate intracellular parasite, Encephalitozoon cuniculi, with or without prior colonic lavages. Although chronic administration of enemas seems to interfere to some degree with the intestinal translocation of the parasite, systemic infection was observed in both manipulated and nonmanipulated animals. The animals responded with antibodies of immunoglobulin A (IgA) and IgG isotypes, reflecting the route of infection. They also produced significant amounts of circulating immune complexes composed of IgA and IgG antibodies and E. cuniculi antigens. Lesions compatible with encephalitozoonosis were seen in the liver, kidney, lung, and brain. In all instances, nonmanipulated animals had more severe lesions than manipulated rabbits given the same dose of parasites. Levels of serum antibodies, circulating immune complexes, and histopathologic changes were associated with the infection dose. The presented data suggest that human microsporidiosis may also be transmitted via the rectal route. It is, therefore, of clinical relevance in view of several reports of microsporidian infections in patients with acquired immunodeficiency

Cellular Basis for Response Diversity in the Olfactory Periphery

An emerging idea in olfaction is that temporal coding of odor specificity can be intrinsic to the primary olfactory receptor neurons (ORNs). As a first step towards understanding whether lobster ORNs are capable of generating odor-specific temporal activity and what mechanisms underlie any such heterogeneity in discharge pattern, we characterized different patterns of activity in lobster ORNs individually and ensemble using patch-clamp recording and calcium imaging. We demonstrate that lobster ORNs show tonic excitation, tonic inhibition, phaso-tonic excitation, and bursting, and that these patterns are faithfully reflected in the calcium signal. We then demonstrate that the various dynamic patterns of response are inherent in the cells, and that this inherent heterogeneity is largely determined by heterogeneity in the underlying intrinsic conductances

An even pattern of xylan substitution is critical for interaction with cellulose in plant cell walls

Xylan and cellulose are abundant polysaccharides in vascular plants and essential for secondary cell wall strength. Acetate or glucuronic acid decorations are exclusively found on even-numbered residues in most of the glucuronoxylan polymer. It has been proposed that this even-specific positioning of the decorations might permit docking of xylan onto the hydrophilic face of a cellulose microfibril. Consequently, xylan adopts a flattened ribbon-like twofold helical screw conformation when bound to cellulose in the cell wall. Here we show that ESKIMO1/XOAT1/TBL29, a xylan-specific O-acetyltransferase, is necessary for generation of the even pattern of acetyl esters on xylan in Arabidopsis. The reduced acetylation in the esk1 mutant deregulates the position-specific activity of the xylan glucuronosyltransferase GUX1, and so the even pattern of glucuronic acid on the xylan is lost. Solid-state NMR of intact cell walls shows that, without the even-patterned xylan decorations, xylan does not interact normally with cellulose fibrils. We conclude that the even pattern of xylan substitutions seen across vascular plants therefore enables the interaction of xylan with hydrophilic faces of cellulose fibrils, and is essential for development of normal plant secondary cell walls.This work was part supported by the Leverhulme Trust grant for the Centre for Natural Material Innovation. J.W.-R., J.J.L. and O.M.T. are supported by studentships from Conicyt Chile and the Cambridge Trusts, the BBSRC Doctoral Training Partnership BB/J014540/1, and a BBSRC Novozymes iCASE award (BB/M015432/1) respectively. We thank K. B. Krogh for co-supervision of O.M.T. The UK 850 MHz solid-state NMR Facility used in this research was funded by EPSRC and BBSRC (Contract reference PR140003), as well as the University of Warwick including via part funding through Birmingham Science City Advanced Materials Projects 1 and 2 supported by Advantage West Midlands (AWM) and the European Regional Development Fund (ERDF)

Bio-Benchmarking of Electronic Nose Sensors

BACKGROUND:Electronic noses, E-Noses, are instruments designed to reproduce the performance of animal noses or antennae but generally they cannot match the discriminating power of the biological original and have, therefore, been of limited utility. The manner in which odorant space is sampled is a critical factor in the performance of all noses but so far it has been described in detail only for the fly antenna. METHODOLOGY:Here we describe how a set of metal oxide (MOx) E-Nose sensors, which is the most commonly used type, samples odorant space and compare it with what is known about fly odorant receptors (ORs). PRINCIPAL FINDINGS:Compared with a fly's odorant receptors, MOx sensors from an electronic nose are on average more narrowly tuned but much more highly correlated with each other. A set of insect ORs can therefore sample broader regions of odorant space independently and redundantly than an equivalent number of MOx sensors. The comparison also highlights some important questions about the molecular nature of fly ORs. CONCLUSIONS:The comparative approach generates practical learnings that may be taken up by solid-state physicists or engineers in designing new solid-state electronic nose sensors. It also potentially deepens our understanding of the performance of the biological system

Molecular and Functional Characterization of the Odorant Receptor2 (OR2) in the Tiger Mosquito Aedes albopictus

In mosquitoes, the olfactory system plays a crucial role in many types of behavior, including nectar feeding, host preference selection and oviposition. Aedes albopictus, known also as the tiger mosquito, is an anthropophilic species, which in the last few years, due to its strong ecological plasticity, has spread throughout the world. Although long considered only a secondary vector of viruses, the potential of its vector capacity may constitute a threat to public health. Based on the idea that an improved understanding of the olfactory system of mosquitoes may assist in the development of control methods that interfere with their behavior, we have undertaken a study aimed at characterizing the A. albopictus Odorant Receptors. Here we report the identification, cloning and functional characterization of the AalOR2 ortholog, that represents the first candidate member of the odorant receptor (OR) family of proteins from A. albopictus. AalOR2 is expressed in the larval heads and antennae of adults. Our data indicate that A. albopictus OR2 (AalOR2) shares a high degree of identity with other mosquito OR2 orthologs characterized to date, confirming that OR2 is one of the most conserved mosquito ORs. Our data indicate that AalOR2 is narrowly tuned to indole, and inhibited by (-)-menthone. In agreement with this results, these two compounds elicit two opposite effects on the olfactory-based behavior of A. albopictus larvae, as determined through a larval behavioral assay. In summary, this work has led to the cloning and de-orphaning of the first Odorant Receptor in the tiger mosquito A. albopictus. In future control strategies this receptor may be used as a potential molecular target

Identification of New Agonists and Antagonists of the Insect Odorant Receptor Co-Receptor Subunit

BACKGROUND: Insects detect attractive and aversive chemicals using several families of chemosensory receptors, including the OR family of olfactory receptors, making these receptors appealing targets for the control of insects. Insect ORs are odorant-gated ion channels, comprised of at least one common subunit (the odorant receptor co-receptor subunit, Orco) and at least one variable odorant specificity subunit. Each of the many ORs of an insect species is activated or inhibited by an unique set of odorants that interact with the variable odorant specificity subunits, making the development of OR directed insect control agents complex and laborious. However, several N-,2-substituted triazolothioacetamide compounds (VUAA1, VU0450667 and VU0183254) were recently shown to act directly on the highly conserved Orco subunit, suggesting that broadly active compounds can be developed. We have explored the chemical space around the VUAA1 structure in order to identify new Orco ligands. PRINCIPAL FINDINGS: We screened ORs from several insect species, using heterologous expression in Xenopus oocytes and an electrophysiological assay, with a panel of 22 compounds structurally related to VUAA1. By varying the nitrogen position in the pyridine ring and altering the moieties decorating the phenyl ring, we identified two new agonists and a series of competitive antagonists. Screening smaller compounds, similar to portions of the VUAA1 structure, also yielded competitive antagonists. Importantly, we show that Orco antagonists inhibit odorant activation of ORs from several insect species. Detailed examination of one antagonist demonstrated inhibition to be through a non-competitive mechanism. CONCLUSIONS: A similar pattern of agonist and antagonist sensitivity displayed by Orco subunits from different species suggests a highly conserved binding site structure. The susceptibility to inhibition of odorant activation by Orco antagonism is conserved across disparate insect species, suggesing the ligand binding site on Orco as a promising target for the development of novel, broadly active insect repellants

Characterization of an Enantioselective Odorant Receptor in the Yellow Fever Mosquito Aedes aegypti

Enantiomers differ only in the left or right handedness (chirality) of their orientations and exhibit identical chemical and physical properties. In chemical communication systems, enantiomers can be differentially active at the physiological and behavioral levels. Only recently were enantioselective odorant receptors demonstrated in mammals while their existence in insects has remained hypothetical. Using the two-microelectrode voltage clamp of Xenopus oocytes, we show that the yellow fever mosquito, Aedes aegypti, odorant receptor 8 (AaOR8) acts as a chiral selective receptor for the (R)-(—)-enantiomer of 1-octen-3-ol, which in the presence of other kairomones is an attractant used by blood-sucking insects to locate their hosts. In addition to steric constraints, chain length and degree of unsaturation play important roles in this recognition process. This is the first characterization of an enantioselective odorant receptor in insects and the results demonstrate that an OR alone, without helper proteins, can account for chiral specificity exhibited by olfactory sensory neurons (OSNs)

Sequence Comparisons of Odorant Receptors among Tortricid Moths Reveal Different Rates of Molecular Evolution among Family Members

In insects, odorant receptors detect volatile cues involved in behaviours such as mate recognition, food location and oviposition. We have investigated the evolution of three odorant receptors from five species within the moth genera Ctenopseustis and Planotrotrix, family Tortricidae, which fall into distinct clades within the odorant receptor multigene family. One receptor is the orthologue of the co-receptor Or83b, now known as Orco (OR2), and encodes the obligate ion channel subunit of the receptor complex. In comparison, the other two receptors, OR1 and OR3, are ligand-binding receptor subunits, activated by volatile compounds produced by plants - methyl salicylate and citral, respectively. Rates of sequence evolution at non-synonymous sites were significantly higher in OR1 compared with OR2 and OR3. Within the dataset OR1 contains 109 variable amino acid positions that are distributed evenly across the entire protein including transmembrane helices, loop regions and termini, while OR2 and OR3 contain 18 and 16 variable sites, respectively. OR2 shows a high level of amino acid conservation as expected due to its essential role in odour detection; however we found unexpected differences in the rate of evolution between two ligand-binding odorant receptors, OR1 and OR3. OR3 shows high sequence conservation suggestive of a conserved role in odour reception, whereas the higher rate of evolution observed in OR1, particularly at non-synonymous sites, may be suggestive of relaxed constraint, perhaps associated with the loss of an ancestral role in sex pheromone reception